Preventing relapse after allogeneic hematopoietic cell transplantation (alloHCT) requires novel approaches to identify and augment beneficial GVL alloimmunity. We propose that chronic lymphocytic leukemia (CLL) is the human model disease to achieve these goals and lessons learned focusing on CLL will improve all transplantation outcomes. Studying CLL patients enables sensitive measurement of minimal residual disease (MRD) which has emerged as the most important predictor of CLL disease free survival following alloHCT. Our innovative research proposal employs massively parallel sequencing of the B and T cell receptor genes to sensitively quantify CLL minimal residual disease MRD while identifying the unique T cell clones providing GVL benefit. We have already demonstrated the feasibility of high-throughput sequencing (HTS) the immunoglobulin heavy chain (IGH) for quantifying CLL MRD. In order to identify beneficial donor T cell clones, ex vivo CLL stimulated donor T cell clones will be isolated by flow cytometry and their unique V T cell receptors (TCR) will be determined by HTS. Following graft infusion, we will quantify these marked anti-CLL TCR clonotypes proliferation and persistence in relation to CLL MRD. Our second innovative application of HTS will assess the overall breadth and depth of donor T and B cell reconstitution following alloHCT. In the final aim, we will test the therapeutic benefit of human allogeneic cytokine induced killer cells (alloCIK) that were successfully translated through human phase I studies supported by our current PPG demonstrating feasibility and low GVHD risk. Overall, we hypothesize that impaired post-transplant donor immune reconstitution with limited T and B cell repertoire diversity limits beneficial GVL alloimmunity and associates with disease relapse. In project 4, we will pioneer massively parallel sequencing of B and T cell receptors to test this hypothesis in CLL patients undergoing alloHCT. We will: 1) quantify CLL MRD, 2) identify beneficial GVL T cell responses, 3) quantify overall immune reconstitution.